PROJECT SUMMARY Chronic wounds are a major threat to public health and present as a comorbid complication with major diseases in humans. Although the proper healing of cutaneous wounds requires collective and coordinated behaviors of multiple cell types, a critical step is the recruitment and function of dermal fibroblasts, which are directed to invade the wound by gradients of a chemoattractant, platelet-derived growth factor (PDGF). A handful of biologicals, most notably recombinant PDGF-BB, are currently approved for treatment of wounds; however, the current treatments lack efficacy in accelerating wound healing, and consequently they have not gained traction in the clinic. These disappointing results underscore how poorly the dynamics of wound healing are understood at the tissue scale and the need to connect knowledge of molecular, cellular, and tissue-level processes to inform and predict outcomes of therapeutic strategies aimed at improving the rate and fidelity of wound repair. We have been developing models of fibroblast chemotaxis with consideration of molecular (polarization of signal transduction), supramolecular (assembly of actomyosin structures), cellular (biased cell movement), and tissue-level (wound invasion) dynamics, which span disparate time (seconds to weeks) and spatial (nm to cm) scales. Many challenges remain. First is the lack of a model connecting, in a mechanistic way, signaling and cytoskeletal dynamics to the mechanics of membrane protrusion/retraction at the cell's leading edge; we call this the molecules to motility problem (Aim 1). It is motivated by our recent discoveries that PDGF chemotaxis and migration biased by gradients of extracellular matrix (ECM) density (haptotaxis) are governed by distinct signaling pathways that affect F-actin dynamics and mechanics in different ways. This fundamental difference is tied to the second critical need, which we call the diversity of cues problem (Aim 2). PDGF is only one spatial cue for fibroblast migration, and hence it is paramount to consider the confluence of chemotactic, haptotactic, and durotactic (gradients in mechanical stiffness) cues that coexist in wounds. Preliminary modeling work has implicated an additional form of spatial bias that we propose to explore: the influence of cell shape, or morphotaxis. The third need is to integrate information about the spatial and biological heterogeneity of the wound. Fast-moving macrophages secrete PDGF and are thus focal sources of chemoattractant, and ECM density and stiffness are also expected to vary in space and time. We refer to the relation of macrophage positions and the dynamic organization of ECM in vivo as the heterogeneous milieu problem (Aim 3).